Resveratrol, a polyphenol presents in grapes and wine, displays antioxidant and anti-inflammatory properties and cytoprotective effect in brain pathologies associated to oxidative stress and neurodegeneration. In previous work, we demonstrated that resveratrol exerts neuroglial modulation, improving glial functions, mainly related to glutamate metabolism. Astrocytes are a major class of glial cells and regulate neurotransmitter systems, synaptic processing, energy metabolism and defense against oxidative stress. This study sought to determine the protective effect of resveratrol against hydrogen peroxide (H2O2)-induced cytotoxicity in C6 astrocyte cell line, an astrocytic lineage, on neurochemical parameters and their cellular and biochemical mechanisms. H2O2 exposure increased oxidative-nitrosative stress, iNOS expression, cytokine proinflammatory release (TNFα levels) and mitochondrial membrane potential dysfunction and decreased antioxidant defenses, such as SOD, CAT and creatine kinase activity. Resveratrol strongly prevented C6 cells from H2O2-induced toxicity by modulating glial, oxidative and inflammatory responses. Resveratrol per se increased heme oxygenase 1 (HO1) expression and extracellular GSH content. In addition, HO1 signaling pathway is involved in the protective effect of resveratrol against H2O2-induced oxidative damage in astroglial cells. Taken together, these results show that resveratrol represents an important mechanism for protection of glial cells against oxidative stress.